Shigella species, the causative agents of bacillary dysentery, are unusually infectious. Volunteer studies have shown as few as 10 ingested organisms are sufficient to produce disease. This is particularly remarkable when one considers the fact that the inoculum must pass through the stomach, with a pH of less than 3.0 in order to reach to intestines where entry into and replication within colonic epithelial cells results in dysentery. Enteric pathogens such as Salmonella species and Vibrio cholora require an infectious dose over 10,000-fold greater than that of Shigella in order to cause enteric disease. Shigella is able to survive exposure to pH 2.5 for several hours whereas Salmonella species are extremely acid sensitive. The acid sensitivity of oral Salmonella-based vaccine strains is a major impediment to their utilization. The objectives of this study are to characterize the genetic and physiologic basis of acid resistance in Shigella species and to utilize this information to construct more efficacious oral vaccine strains. We have identified a gene from Shigella flexneri, rpo/s, which confers acid resistance on an acid sensitive Escherichia coli, HB101. Rpo/s encodes a stationary phase sigma factor which acts indirectly in conferring acid resistance on Shigella species. In order to identify specific genes required for acid resistance, 36 acid sensitive mutants were obtained using transposon mutagenesis. Three of these classes of mutations contain insertions in genes with known homologies: PhoN Salmonella typhimurium, MxiA, a plasmid encoded gene required for virulence in S. flexneri, and OmpT, a protease in E. coli. The remaining six classes of insertions fall within genes for which there is no significant DNA homology with known genes. The expression of some of these acid- resistance genes appears to be regulated by rpo/s. Identification and further analysis of this set of acid resistant genes will be useful in the construction of acid resistant oral vaccine strains.